Center for Vision Research and Department of Ophthalmology, State University of New York Upstate Medical University, Syracuse, NY.
Department of Neuroscience and Physiology, State University of New York Upstate Medical University, Syracuse, NY.
J Cell Biol. 2018 Aug 6;217(8):2831-2849. doi: 10.1083/jcb.201711104. Epub 2018 Jun 26.
Physical properties of primary cilia membranes in living cells were examined using two independent, high-spatiotemporal-resolution approaches: fast tracking of single quantum dot-labeled G protein-coupled receptors and a novel two-photon super-resolution fluorescence recovery after photobleaching of protein ensemble. Both approaches demonstrated the cilium membrane to be partitioned into corralled domains spanning 274 ± 20 nm, within which the receptors are transiently confined for 0.71 ± 0.09 s. The mean membrane diffusion coefficient within the corrals, = 2.9 ± 0.41 µm/s, showed that the ciliary membranes were among the most fluid encountered. At longer times, the apparent membrane diffusion coefficient, = 0.23 ± 0.05 µm/s, showed that corral boundaries impeded receptor diffusion 13-fold. Mathematical simulations predict the probability of G protein-coupled receptors crossing corral boundaries to be 1 in 472. Remarkably, latrunculin A, cytochalasin D, and jasplakinolide treatments altered the corral permeability. Ciliary membranes are thus partitioned into highly fluid membrane nanodomains that are delimited by filamentous actin.
使用两种独立的、高时空分辨率方法研究了活细胞中初级纤毛膜的物理性质：快速跟踪单个量子点标记的 G 蛋白偶联受体和一种新的双光子超分辨率荧光漂白后蛋白集合的恢复。这两种方法都表明纤毛膜被分隔成 274 ± 20nm 的围栏域，其中受体被短暂限制在 0.71 ± 0.09s 内。在围栏内的平均膜扩散系数， = 2.9 ± 0.41 µm/s，表明纤毛膜是遇到的最具流动性的膜之一。在更长的时间内，表观膜扩散系数， = 0.23 ± 0.05 µm/s，表明围栏边界使受体扩散受阻 13 倍。数学模拟预测 G 蛋白偶联受体穿过围栏边界的概率为 1 比 472。值得注意的是，Latrunculin A、细胞松弛素 D 和 Jasplakinolide 处理改变了围栏的渗透性。因此，纤毛膜被分隔成高度流动的膜纳米域，这些域由丝状肌动蛋白限定。
